Lipogenesis is regulated by modulating (a) synthesis of its component enzymes and (b) catalytic efficiency of the pace-setting enzyme of the pathway. Co-ordinate changes in the catalytic efficiency and synthesis of the lipogenic enzymes in vivo are caused by developmental and nutritional stimuli and in culture by hormones and free fatty acids. Our objective is to understand the molecular basis for the co-ordinate regulation of enzyme synthesis and overall pathway function. In the liver cell culture system which we have developed thyroid hormone and insulin stimulate and glucagon inhibits activity of the fatty acid synthesis pathway and synthesis of one or more of the lipogenic enzymes. These hormone effects are large, rapid, specific and elicited by physiological concentrations, making the system ideal for the study of the mechanisms of action of thyroid hormone and glucagon. We have purified and prepared specific antibodies against two of the lipogenic enzymes, malic enzyme, and fatty acid synthetase. Using immunological techniques, we propose to develop assays for the specific mRNAs for each of these enzymes. The assays will be used to estimate the level of translatable mRNA present in cells under different hormonal and nutritional conditions and to monitor purification of the mRNAs. Using recombinant DNA techniques we propose to isolate cloned cDNA or genomic DNA sequences for fatty acid synthetase and malic enzyme. These cloned sequences will be used as probes to study the hormonal regulation of the concentration, synthesis and degradation of fatty acid synthetase and malic enzyme mRNAs in liver cells in culture.